The antinociception of gestation (GSA) and its hormonal simulation (HSPA) result from the integration of multiple components and requires concomitant activity of dynorphin (Dyn)/K and enkephalin/6 opioid receptor (enk/DOR) spinal analgesic systems. Enhanced release of spinal Dyn during HSPA results from the loss of inhibition of evoked Dyn release by nociceptin and the conversion of its DOR-coupled inhibition to facilitation. Afferent input via the hypogastric nerve (HGN) and activation of spinal noradrenergic receptors (NARs) are also required for GSA and HSPA. However, the mechanism(s) underlying these changes remains unknown. Our biochemical and preliminary immunohistochemical (IHC) data have allowed us to formulate an integrated model of the neuronal interactions that underlie GSA and HSPA. We hypothesize that the shift from inhibition to facilitation of enk release by DOR is triggered by increased release of spinal vasoactive intestinal polypeptide (VIP) from varicosities that appose Dyn neurons. This, alone or in combination with activation of DORs that are expressed by Dyn neurons, augments their cAMP content. We contend that these events are critical for the loss of DOR- coupled inhibition and the appearance of DOR-coupled facilitation of Dyn release as well as for the loss of its inhibition via nociceptin. We also postulate that during HSP, there are enhanced Dyn releasing effects of the spinal a2c-NARs that are expressed by Dyn neurons. Four major specific aims are proposed with which to validate this model: (1) Determine the effect of pregnancy and HSP on the release of spinal VIP. This will be assessed using ex vivo spinal tissue via radioimmunoassay as well as via in vivo VIP receptor internalization visualized by IHC. (2) Determine (a) the effect of VIP receptor activation, alone and in combination with DOR activation, on the regulation of spinal Dyn release from control spinal tissue and (b) determine the effects of a2c-NAR agonists and antagonists on Dyn release from spinal tissue obtained from control and HSP animals. (3) Determine the consequences to GSA and HSPA of in vivo (a) spinal VIP receptor blockade and (b) inhibition of spinal protein kinase A. (4) Determine the spatial relationships among components of GSA and HSPA and the influence of ovarian sex steroids on their prevalence. Activation of primary afferents would be expected to release excitatory transmitters that should produce pain, not ameliorate its perception. However, there are data indicating that primary afferent activity can also enhance opioid antinociception. Our proposed experiments should provide a framework for further understanding this paradox. The proposed research should also shed light on the ability of ovarian sex steroids to alter (visceral) injury-induced spinal remodeling to produce hypoalgesia instead of the neuropathic pain that often results during the nonpregnant state.